Transcript Slide 1
Strategies for detection of melamine
in milk products
Xin Wu
Literature Seminar
05/03/2010
Objective
Compare two methods to detect of melamine in
milk products.
Melamine
2,4,6-triamino-s-triazine, MM
In manufacture of:
Plastics,
Flame retardant
Fertilizer
Toxicity
Acute toxicity
Chronic toxicity
Contaminants in
many kinds of food
http://www.toothpastefordinner.co
m/archives/2008/Dec/
Summary of the detection methods
Nitrogen methods
Kjeldahl method
Most traditional
subtractive method
veracity is poor
Other methods
sublimation and picric acid
weight methods
high veracity, complicated operation,
time-consuming
GC with UV/MS detection
Expensive, derivatization before GC
.sensitive
LC-MS/MS
LOD low, sensitive but complicated
Kjeldahl method
Degradation: Protein + H2SO4 → (NH4)2SO4(aq) + CO2(g)
+ SO2(g) + H2O(g)
Liberation of ammonia: (NH4)2SO4(aq) + 2NaOH →
Na2SO4(aq) + 2H2O(l) + 2NH3(g)
Capture of ammonia: B(OH)3 + H2O + NH3 → NH4+ +
B(OH)4–
Back-titration: B(OH)3 + H2O + Na2CO3 → NaHCO3(aq)
+ NaB(OH)4(aq) + CO2(g) + H2O
Paper I
Residue analysis of melamine in milk Products
by micellar electrokinetic capillary
chromatography with amperometric detection
Jinyan Wang, Lianmei Jiang, Qingcui Chu , Jiannong Ye
Food Chemistry 121 (2010) 215–219
Capillary electrophoresis (CE)
Separate ionic species by their charge and mass.
UV or fluorescence detection
Advantages of CE over HPLC
Micellar electrokinetic chromatography
Modification of capillary electrophoresis
Sample separated between micelles and buffer
C(Surfactant) >CMC
in open capillaries under
alkaline conditions
Sodium dodecyl sulfate
(SDS)
Shegeru,Japan Chemical Journal 8:291-301
Materials and Preparation
laboratory-built CE–AD system
MM ; Milk and infant formula milk powder samples;
running buffer (8 mmol/L SDS/20 mmol/L H3BO3–
Na2B4O7 solution)
Weighed-Extracted by ultrasonic bath-filtered-CEAD system
Effect of the potential applied to the
working electrode
1 Above 750 mV, oxidation
current increased rapidly
with applied
potential.
2 Above 1200 mV, unstable
baseline
3 1150mV is best choice
Effects of the pH value
1 pH range 7.4-9.2
2 MM always overlapped
with the unknown
compound peak in real
samples
3 pH is optimum pH
Effects of SDS
1 at a fixed pH value 7.4
2 longer retention time
with increased
concentration of SDS
3 8 mmol/L SDS is
optimum concentratioin
Electropherograms
Recovery of samples
Conclusion
A new method of MECC-AD to residue analysis of
melamine
For the real world sample
Run time is 9 min
Good recoveries ranging from 83.3% to 105.5% of MM
were determined
Practical method with sample preconcentration
Paper II
A sensitive and validated method for
determination of melamine residue in liquid
milk by reversed phase high-performance liquid
Chromatography with solid-phase extraction
Hanwen Sun, Lixin Wang, Lianfeng Ai, Shuxuan Liang, Hong Wu
Food Control 21 (2010) 686–691
HPLC
separate, identify, and quantify compounds based on their
idiosyncratic polarities and interactions with the column's
stationary phase
A pump propel the mobile
phase and analyte through
densely packed column
Widely used
Various types of HPLC
http://en.wikipedia.org/wiki/File:Hplc.JPG
solid-phase extraction (SPE)
Concentrate and purify samples for analysis
Used to separate analytes from a variety of matrices:
(1) blood
(2) animal tissue
(3) water etc.
Mobile phase (liquid and analyte); Stationary Phase (solid)
Silica
Materials and Methods
Cleanert PCX-SPE cartridges (3 mL/60 mg), LC–MS/MS
(Thermo Electron Corp., San Jose, CA, USA), an SeQuant
ZIC HILIC column.
Sodium n-heptanesulfonate, acetonitrile, melamine, 1%
trichloroacetic acid, 2.2% lead acetate solution, liquid milk
sample
Confirmation by LC-MS/MS, Sample extraction and clean-
up, HPLC analysis,
The effects of pH
pH =2.7 is the
optimum value
The effects of the ratio (v/v) between 10 mM
sodium n-heptanesulfate (pH 2.7) and acetonitrile
retention time
of melamine
shorted with
the decrease of
ratio value.
The LOD and recovery of sample
Conclusion
A new method of RP-HPLC-SPE to residue analysis of
melamine
Real world sample
Run time is 10 min
Good recoveries ranging from 85.5–99.3%of MM were
determined
Comparison of the two methods
TIME(min)
LOD (10-6g/ml)
Recovery
MECC-AD
9
2.1
83.3% to 105.5%
RP-HPLC-SPE
10
3
85.5–99.3%
Acknowledgements
Professor Kermit K. Murray
Murray Research Group
Audience
Any questions?